The sheer number of deaths attributable to human immunodeficiency virus (HIV)-induced acquired immunodeficiency syndrome (AIDS) is sobering, making the development of a preventative vaccine all the more urgent. Historically, the most effective vaccines are live-attenuated or inactivated pathogens. Safety concerns exclude these strategies in the case of HIV. In that regard both adenoviruses (Ad) that replicate (Ad?E3) and those that do not (Ad?E1?E3), represent a viable vaccine delivery method. In a recent comparison however, replicating Ad at lower doses induced better HIV transgene-specific immune responses than the non-replicating vector. Since the two vectors have the same surface proteins the advantage is most likely derived from virus-host cell interactions perpetuated by products of the viral genes that are severely muted in cells infected by the non-replicating vector. These virus-host cell interactions enhance transgene- specific immune responses and are herein termed the adjuvant quality of replicating Ad vectors. The goal of this proposed research is to develop a better understanding of the factors that influence the immune responses to the transgene within replicating Ad vaccine vectors. We test the global hypothesis that the adjuvant quality of replicating Ad is dependent on the products of the virus E1 and E4 genes. A rather curious discovery is that endoreduplication, a phenomenon that associates with DNA damage and genetic instability often observed in cancer, occurs in all Ad-infected cell types. Therefore the virus-host cell interactions that promote endoreduplication may be important to the virus pathology. If we can define the viral and cellular factors involved in this process we will improve our understanding of both adenoviral biology and cancer. Most importantly for replicating Ad-HIV vaccine vector is that, as in other systems, here too DNA damage/endoreduplication may also engage the immune system. Using biochemical, molecular biology, microscopy and flow cytometry techniques I will make and characterize various cell lines and gp120 transgene-bearing E1B55K/E3-deleted viruses additionally deleted of E4. These will be used in aim 1 to define the mechanism by which Ad induces endoreduplication in infected cells. In aim 2 I will determine if viruses that induce differing levels of endoreduplicaton elicit differing levels of Ad- and/or gp120-specific immune responses in immunized mice. The results of this study will inform the design and development of more effective replicating Ad-HIV vaccines.